Dissecting ITC data of metal ions binding to ligands and proteins

Johnson, Rachel A.; Manley, Olivia M.; Spuches, Anne M.; Grossoehme, Nicholas E.

doi:10.1016/j.bbagen.2015.08.018

Cited by 34 publications

(28 citation statements)

References 70 publications

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“…We conducted relative comparisons of the divalent metal interaction with isolated RNH probed by ITC and CD. Although a buffer effect may contribute to the absolute binding thermodynamics [20][21][22][23], specific thermal and structural features of Zn 2+ -binding were obtained, consistent with a previous study by DeStefano's group [8]. Further, Mg 2+ binding thermodynamics to RNH were determined using conditions similar to those applied in previous NMR studies.…”

Section: Discussionsupporting

confidence: 84%

“…Metal ions play important roles in molecular interactions and functions in biology: their thermodynamics studies have provided critical insights into the molecular mechanism [1][2][3][4][5]. Human immunodeficiency virus-1 reverse transcriptase (HIV-RT) possesses both nucleotide polymerization and ribonuclease H (RNH) activities, the latter of which degrades RNA of an RNA-DNA hybrid.…”

Section: Introductionmentioning

confidence: 99%

“…In this study, we analyzed the binding thermodynamics of Mn 2+ , Zn 2+ , and Mg 2+ to isolated RNH using ITC. More specifically, we carried out a relative comparison of these metal ions in unique experimental conditions, i.e., thermodynamically detected metal-protein interaction in a background that metal-buffer interaction may exist [20][21][22][23]. In contrast to the previous study for direct binding of Mg 2+ to RT [7], little heat was observed for direct binding to isolated RNH, presumably due to its weak binding affinity.…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Binding thermodynamics of metal ions to HIV-1 ribonuclease H domain

Oda

Zhang

Inaba

et al. 2018

J Therm Anal Calorim

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Metal-protein interactions are not necessarily tight in many transient biological processes, such as cellular signaling, enzyme regulation, and molecular recognition. Here, we analyzed the binding thermodynamics and characterized the structural effect of divalent metal ions, i.e. Mn 2+ , Zn 2+ , and Mg 2+ , to the isolated ribonuclease H (RNH) of human immunodeficiency virus (HIV) using isothermal titration calorimetry (ITC) and circular dichroism. The binding thermodynamics of Mg 2+ to RNH was determined using competition ITC experiments, and the binding affinity of Mg 2+ was found to be about 40-and 400-times lower than those of Mn 2+ and of Zn 2+ , respectively. The structural analysis showed that Mg 2+ binding had little effect on the thermal stability of RNH, while Zn 2+ and Mn 2+ binding increased the stability. The thermodynamic characteristics of RNH metal binding, compared to intact HIV reverse transcriptase, and a possible mechanism of conformational change induced upon metal ion binding, in correlation with the structure-function relationship, are discussed.

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Section: Discussionsupporting

confidence: 84%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Binding thermodynamics of metal ions to HIV-1 ribonuclease H domain

Oda

Zhang

Inaba

et al. 2018

J Therm Anal Calorim

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“…It should be noted that same enthalpies of the formation of Zn(Hk) 2 were obtained by both titrations of Zn 2+ ions into peptide and titrations of peptide into Zn 2+ (Figs S16 and S17). Because Zn 2+ complexation by cysteine-containing peptides is accompanied by thiol deprotonation, the experimental enthalpy (Δ H ITC ) must be corrected for the heat of protonation of the buffer4546, as indicated in equation (4):…”

Section: Resultsmentioning

confidence: 99%

Metal-coupled folding as the driving force for the extreme stability of Rad50 zinc hook dimer assembly

Kochańczyk

Nowakowski

Wojewska

et al. 2016

Sci Rep

125

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The binding of metal ions at the interface of protein complexes presents a unique and poorly understood mechanism of molecular assembly. A remarkable example is the Rad50 zinc hook domain, which is highly conserved and facilitates the Zn2+-mediated homodimerization of Rad50 proteins. Here, we present a detailed analysis of the structural and thermodynamic effects governing the formation and stability (logK12 = 20.74) of this evolutionarily conserved protein assembly. We have dissected the determinants of the stability contributed by the small β-hairpin of the domain surrounding the zinc binding motif and the coiled-coiled regions using peptides of various lengths from 4 to 45 amino acid residues, alanine substitutions and peptide bond-to-ester perturbations. In the studied series of peptides, an >650 000-fold increase of the formation constant of the dimeric complex arises from favorable enthalpy because of the increased acidity of the cysteine thiols in metal-free form and the structural properties of the dimer. The dependence of the enthalpy on the domain fragment length is partially compensated by the entropic penalty of domain folding, indicating enthalpy-entropy compensation. This study facilitates understanding of the metal-mediated protein-protein interactions in which the metal ion is critical for the tight association of protein subunits.

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“…In order to achieve the outlined research goals, we decided to apply isothermal titration calorimetry (ITC) as a unique method, which can be used for studies of the interactions of spectroscopically and magnetically silent metal ions such as Ca 2+ , providing complete thermodynamic profile of a reaction (Δ G ITC , Δ H 0 ITC , and Δ S 0 ITC ), in which bio‐molecule participates. ITC makes a contribution to a basic knowledge about the nature of forces, which are involved in those interactions.…”

Section: Introductionmentioning

confidence: 99%

Thermodynamics of the Interactions of Aminobisphosphonates and Their Calcium Complexes with Bovine Serum Albumin

Gałęzowska

Chmielewska

2018

Chemistry & Biodiversity

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Binding of bisphosphonates (BPs) to plasma proteins was investigated in the 1990s as a pharmacokinetic issue in order to fully understand bio-distribution of BP drugs which are successfully used for the treatment of several bone-related diseases. It has been hypothesized that binding to these proteins occurs with low to moderate affinity despite of unfavorable hydrophilicity of BPs, and Ca was identified as a strong catalyst of this binding. However, these studies mainly consisted in the separation and quantification of bound and unbound drug or protein fractions using chromatographic techniques without an outcome on the molecular level. Presented thermodynamic studies analyze the interactions of three N-BPs as well as their Ca complexes with bovine serum albumine (BSA) by means of isothermal calorimetry. The studies reveal spontaneous enthalpy favored interactions of N-BPs (amino-containing BPs) with BSA, which are enhanced by the presence of Ca ions up to ~15-fold, strongly depending on N-BP. Those are low affinity binding events, comparable to Ca -N-BP interactions, which most likely occur at Ca binding site(s). It is a first example of estimation of thermodynamic forces of interactions of free and calcium-bound N-BPs with albumin.

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Dissecting ITC data of metal ions binding to ligands and proteins

Cited by 34 publications

References 70 publications

Binding thermodynamics of metal ions to HIV-1 ribonuclease H domain

Binding thermodynamics of metal ions to HIV-1 ribonuclease H domain

Metal-coupled folding as the driving force for the extreme stability of Rad50 zinc hook dimer assembly

Thermodynamics of the Interactions of Aminobisphosphonates and Their Calcium Complexes with Bovine Serum Albumin

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